多药耐药转运蛋白Snq2p和Pdr5p介导酿酒酵母中的咖啡因外排。

Multidrug resistance transporters Snq2p and Pdr5p mediate caffeine efflux in Saccharomyces cerevisiae.

作者信息

Tsujimoto Yoshiyuki, Shimizu Yoshihiro, Otake Kazuya, Nakamura Tatsuya, Okada Ryutaro, Miyazaki Toshitaka, Watanabe Kunihiko

机构信息

a Graduate School of Life and Environmental Sciences , Kyoto Prefectural University , Kyoto , Japan.

出版信息

Biosci Biotechnol Biochem. 2015;79(7):1103-10. doi: 10.1080/09168451.2015.1010476. Epub 2015 Feb 16.

DOI:10.1080/09168451.2015.1010476

PMID:25686090

Abstract

SNQ2 was identified as a caffeine-resistance gene by screening a genomic library of Saccharomyces cerevisiae in a multicopy vector YEp24. SNQ2 encodes an ATP-binding cassette transporter and is highly homologous to PDR5. Multicopy of PDR5 also conferred resistance to caffeine, while its resistance was smaller than that of SNQ2. Residual caffeine contents were analyzed after transiently exposing cells to caffeine. The ratios of caffeine contents were 21.3 ± 8.8% (YEp24-SNQ2) and 81.9 ± 8.7% (YEp24-PDR5) relative to control (YEp24, 100%). In addition, multicopies of SNQ2 or PDR5 conferred resistance to rhodamine 6G (R6G), which was widely used as a substrate for transport assay. R6G was exported by both transporters, and their efflux activities were inhibited by caffeine with half-maximal inhibitory concentrations of 5.3 ± 1.9 (YEp24-SNQ2) and 17.2 ± 9.6 mM (YEp24-PDR5). These results demonstrate that Snq2p is a more functional transporter of caffeine than Pdr5p in yeast cells.

摘要

通过在多拷贝载体YEp24中筛选酿酒酵母的基因组文库，SNQ2被鉴定为一种咖啡因抗性基因。SNQ2编码一种ATP结合盒转运蛋白，与PDR5高度同源。PDR5的多拷贝也赋予了对咖啡因的抗性，但其抗性小于SNQ2。在将细胞短暂暴露于咖啡因后，分析了残余咖啡因含量。相对于对照（YEp24，100%），咖啡因含量的比率分别为21.3±8.8%（YEp24-SNQ2）和81.9±8.7%（YEp24-PDR5）。此外，SNQ2或PDR5的多拷贝赋予了对罗丹明6G（R6G）的抗性，R6G被广泛用作转运测定的底物。两种转运蛋白都能输出R6G，并且它们的外排活性受到咖啡因的抑制，其半数最大抑制浓度分别为5.3±1.9（YEp24-SNQ2）和17.2±9.6 mM（YEp24-PDR5）。这些结果表明，在酵母细胞中，Snq2p是一种比Pdr5p更具功能的咖啡因转运蛋白。